The broad long-term overall goal of the proposed research is to understand the physiological role of estrogen (E2) and estrogen receptors (ERs) during neuronal development and to determine to what extent environmental estrogens (EEs) can mimic or inhibit the effects of E2 in these developing neurons. Mechanisms of rapid E2-mediated signal transduction are not well understood. These actions vary greatly from cell type to cell type, and may even vary in the same cell. In addition to reproductive tissues &breast cancers, cells of the nervous system are a clinically very important E2 targets. Environmental estrogens, also known as endocrine disrupting chemicals, are a diverse group of compounds that can mimic or antagonize the normal actions of E2. The extent to which EEs impact the developing nervous system is unclear. Proposed experiments using primary cultures of estrogen-sensitive neonatal rat cerebellar neurons and the developing cerebellum as non-sexually related neuronal models, will address the following Specific Aims - Specific Aim 1 is to determine whether or not structurally diverse EEs modulate ERK1/2 signaling in developing cerebellar neurons. It is hypothesized that in E2-responsive neurons, EEs rapidly modulate ERK1/2 signaling and neuronal physiology in an E2-like fashion, and that differences in the chemical structure of individual EEs determine their ability to activate ERK-signaling &/or antagonize the rapid actions of E2. Specific Aim 2 is to determine the nature of the signaling mechanism underlying E2-mediated rapid activation of ERK-signaling. It is hypothesized that a plasma membrane, localized version of ERp is acting as the mediator of rapid E2/EE-induced ERKsignaling. Membrane E2-binding proteins from primary granule cell and hippocampal neuronal cultures will be affinity purified and identified through LC-tandem mass spectrometry. Immunocytochemical and loss-of-function experiments are proposed to demonstrate the function of identified candidate membrane ERs. The ability of ERa, ERp, and the orphan G-protein coupled receptor GPR30 to act as the membrane ER will be directly addressed.